Methods of treating biofilms

ABSTRACT

A method of treating or preventing a condition caused by a biofilm in a subject is carried out by: providing a subject having, or susceptible to, a condition mediated by a biofilm produced by a microorganism, where the biofilm comprises a matrix and the microorganism on a surface and administering to the subject an active agent under conditions effective for the condition caused by a biofilm in the subject to be treated or prevented, where the active agent is a compound of general Formula (I): or a pharmaceutically acceptable salt thereof.

RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Patent Application Ser. No. 61/872,911, filed Sep. 3, 2013, the disclosure of which is incorporated herein by reference in its entirety.

BACKGROUND OF THE INVENTION

Microbial biofilms in infections and in industrial systems present significant problems due to their recalcitrance to effective treatment. See, e.g., U.S. Pat. No. 8,513,305 to Davies et al. Accordingly, there is a need for new ways to combat or disrupt biofilm formation, both in vivo and ex vivo.

SUMMARY OF THE INVENTION

Described herein is a method of treating, inhibiting or preventing a condition caused by a biofilm in a subject, the method comprising: providing a subject having, or susceptible to, a condition mediated by a biofilm produced by a microorganism, where the biofilm comprises a matrix and the microorganism on a surface and administering to the subject an active agent as described herein under conditions effective for the condition caused by a biofilm in the subject to be treated, inhibited or prevented.

Also described herein is a method of treating or inhibiting formation of a biofilm on a surface, the method comprising: providing a surface having or being susceptible to formation of a biofilm produced by a microorganism, where the biofilm comprises a matrix and the microorganism on the surface, and administering to the surface an active agent as described herein under conditions effective for formation of the biofilm on the surface to be treated or inhibited.

A still further aspect of the present invention is an active agent as described herein for use in a method of treatment as described herein, or for use in the preparation of a medicament for carrying out a method of treatment as described herein.

The present invention is explained in greater detail in the specification set forth below. The disclosures of all United States Patents cited herein are incorporated herein by reference.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1. In vitro biofilm intensity with and without treatment with of the SOD mimetic MnTE-2-PyP.

FIG. 2. Bacterial Counts from femur tissues. Keflex with MnTE compared to Keflex alone showed significant decrease in bacteria counts (p=0.007).

FIG. 3. Fracture Healing in Infected Fracture Model of Blofilm forming Staphylococcus Aureus. Little or no fracture healing was seen in the group of animals with no treatment. Antibiotic and Antibiotic plus SOD mimetic showed significantly improved fracture healing compared to no treatment. Antibiotic with mimetic was significantly better than antibiotic alone (p=0.001) for fracture healing at 2 wks.

DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS

The present invention is primarily concerned with the treatment of human subjects, but the invention may also be carried out on animal subjects, particularly mammalian subjects such as dogs, cats, livestock and horses for veterinary purposes. While subjects may be of any suitable age, the subjects are in some embodiments neonatal, infant, juvenile, adolescent, adult, or geriatric subjects.

“Treat” as used herein refers to any type of treatment that imparts a benefit to a patient or subject matter as described herein, particularly delaying or retarding the onset or progression of the conditions described herein, or reducing the severity of symptoms, or speeding or improving recovery therefrom.

“Pharmaceutically acceptable” as used herein means that the compound or composition is suitable for administration to a subject to achieve the treatments described herein, without unduly deleterious side effects in light of the severity of the disease and necessity of the treatment.

“Combination” as used herein with respect to a method of administration (e.g., an active compound and an antibiotic administered in combination) includes administering the the two or more compounds simultaneously, or sequentially, sufficiently close in time to produce a combined therapeutic or treatment effect.

“Alkyl” as used herein alone or as part of another group, refers to a straight or branched chain hydrocarbon containing from 1 to 10 carbon atoms. Representative examples of alkyl include, but are not limited to, methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, 3-methylhexyl, 2,2-dimethylpentyl, 2,3-dimethylpentyl, n-heptyl, n-octyl, n-nonyl, n-decyl, and the like. “Lower alkyl” as used herein, is a subset of alkyl, in some embodiments preferred, and refers to a straight or branched chain hydrocarbon group containing from 1 to 4 carbon atoms. Representative examples of lower alkyl include, but are not limited to, methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, tert-butyl, and the like. The term “akyl” or “loweralkyl” is intended to include both substituted and unsubstituted alkyl or loweralkyl unless otherwise indicated and these groups may be substituted with groups selected from halo (e.g., haloalkyl), alkyl, haloalkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, heterocyclo, heterocycloalkyl, hydroxyl, alkoxy (thereby creating a polyalkoxy such as polyethylene glycol), alkenyloxy, alkynyloxy, haloalkoxy, cycloalkoxy, cycloalkylalkyloxy, aryloxy, arylalkyloxy, heterocyclooxy, heterocyclolalkyloxy, mercapto, alkyl-S(O)_(m), haloalkyl-S(O)_(m), alkenyl-S(O)_(m), alkynyl-S(O)_(m) cycloalkyl-S(O)_(m), cycloalkylalkyl-S(O)_(m), aryl-S(O)_(m), arylalkyl-S(O)_(m), heterocyclo-S(O)_(m), heterocycloalkyl-S(O)_(m), amino, carboxy, alkylamino, alkenylamino, alkynylamino, haloalkylamino, cycloalkylamino, cycloalkylalkylamino, arylamino, arylalkylamino, heterocycloamino, heterocycloalkylamino, disubstituted-amino, acylamino, acyloxy, ester, amide, sulfonamide, urea, alkoxyacylamino, aminoacyloxy, nitro or cyano where m=0, 1, 2 or 3.

“Alkenyl” as used herein alone or as part of another group, refers to a straight or branched chain hydrocarbon containing from 1 to 10 carbon atoms (or in loweralkenyl 1 to 4 carbon atoms) which include 1 to 4 double bonds in the normal chain. Representative examples of alkenyl include, but are not limited to, vinyl, 2-propenyl, 3-butenyl, 2-butenyl, 4-pentenyl, 3-pentenyl, 2-hexenyl, 3-hexenyl, 2,4-heptadiene, and the like. The term “alkenyl” or “loweralkenyl” is intended to include both substituted and unsubstituted alkenyl or loweralkenyl unless otherwise indicated and these groups may be substituted with groups as described in connection with alkyl and loweralkyl above.

“Alkynyl” as used herein alone or as part of another group, refers to a straight or branched chain hydrocarbon containing from 1 to 10 carbon atoms (or in loweralkynyl 1 to 4 carbon atoms) which include 1 triple bond in the normal chain. Representative examples of alkynyl include, but are not limited to, 2-propynyl, 3-butynyl, 2-butynyl, 4-pentynyl, 3-pentynyl, and the like. The term “alkynyl” or “loweralkynyl” is intended to include both substituted and unsubstituted alkynyl or loweralknynyl unless otherwise indicated and these groups may be substituted with the same groups as set forth in connection with alkyl and loweralkyl above.

“Cycloalkyl” as used herein alone or as part of another group, refers to a saturated or partially unsaturated cyclic hydrocarbon group containing from 3, 4 or 5 to 6, 7 or 8 carbons (which carbons may be replaced in a heterocyclic group as discussed below). Representative examples of cycloalkyl include, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. These rings may be optionally substituted with additional substituents as described herein such as halo or loweralkyl. The term “cycloalkyl” is generic and intended to include heterocyclic groups as discussed below unless specified otherwise.

“Heterocyclic group” or “heterocyclo” as used herein alone or as part of another group, refers to an aliphatic (e.g., fully or partially saturated heterocyclo) or aromatic (e.g., heteroaryl) monocyclic- or a bicyclic-ring system. Monocyclic ring systems are exemplified by any 5 or 6 membered ring containing 1, 2, 3, or 4 heteroatoms independently selected from oxygen, nitrogen and sulfur. The 5 membered ring has from 0-2 double bonds and the 6 membered ring has from 0-3 double bonds. Representative examples of monocyclic ring systems include, but are not limited to, azetidine, azepine, aziridine, diazepine, 1,3-dioxolane, dioxane, dithiane, furan, imidazole, imidazoline, imidazolidine, isothiazole, isothiazoline, isothiazolidine, isoxazole, isoxazoline, isoxazolidine, morpholine, oxadiazole, oxadiazoline, oxadiazolidine, oxazole, oxazoline, oxazolidine, piperazine, piperidine, pyran, pyrazine, pyrazole, pyrazoline, pyrazolidine, pyridine, pyrimidine, pyridazine, pyrrole, pyrroline, pyrrolidine, tetrahydrofuran, tetrahydrothiophene, tetrazine, tetrazole, thiadiazole, thiadiazoline, thiadiazolidine, thiazole, thiazoline, thiazolidine, thiophene, thiomorpholine, thiomorpholine sulfone, thiopyran, triazine, triazole, trithiane, and the like. Bicyclic ring systems are exemplified by any of the above monocyclic ring systems fused to an aryl group as defined herein, a cycloalkyl group as defined herein, or another monocyclic ring system as defined herein. Representative examples of bicyclic ring systems include but are not limited to, for example, benzimidazole, benzothiazole, benzothiadiazole, benzothiophene, benzoxadiazole, benzoxazole, benzofuran, benzopyran, benzothiopyran, benzodioxine, 1,3-benzodioxole, cinnoline, indazole, indole, indoline, indolizine, naphthyridine, isobenzofuran, isobenzothiophene, isoindole, isoindoline, isoquinoline, phthalazine, purine, pyranopyridine, quinoline, quinolizine, quinoxaline, quinazoline, tetrhydroisoquinoline, tetrahydroquinoline, thiopyranopyridine, and the like. These rings include quaternized derivatives thereof and may be optionally substituted with groups selected from halo, alkyl, haloalkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, heterocyclo, heterocycloalkyl, hydroxyl, alkoxy, alkenyloxy, alkynyloxy, haloalkoxy, cycloalkoxy, cycloalkylalkyloxy, aryloxy, arylalkyloxy, heterocyclooxy, heterocyclolalkyloxy, mercapto, alkyl-S(O)_(m), haloalkyl-S(O)_(m), alkenyl-S(O)_(m), alkynyl-S(O)_(m), cycloalkyl-S(O)_(m), cycloalkylalkyl-S(O)_(m), aryl-S(O)_(m), arylalkyl-S(O)_(m), heterocyclo-S(O)_(m), heterocycloalkyl-S(O)_(m), amino, alkylamino, alkenylamino, alkynylamino, haloalkylamino, cycloalkylamino, cycloalkylalkylamino, arylamino, arylalkylamino, heterocycloamino, heterocycloalkylamino, disubstituted-amino, acylamino, acyloxy, ester, amide, sulfonamide, urea, alkoxyacylamino, aminoacyloxy, nitro or cyano where m=0, 1, 2 or 3.

“Aryl” as used herein alone or as part of another group, refers to a monocyclic carbocyclic ring system or a bicyclic carbocyclic fused ring system having one or more aromatic rings. Representative examples of aryl include, azulenyl, indanyl, indenyl, naphthyl, phenyl, tetrahydronaphthyl, and the like. The term “aryl” is intended to include both substituted and unsubstituted aryl unless otherwise indicated and these groups may be substituted with the same groups as set forth in connection with alkyl and loweralkyl above.

“Arylalkyl” as used herein alone or as part of another group, refers to an aryl group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples of arylalkyl include, but are not limited to, benzyl, 2-phenylethyl, 3-phenylpropyl, 2-naphth-2-ylethyl, and the like.

“Heteroaryl” as used herein is as described in connection with heterocyclo above.

“Alkoxy” as used herein alone or as part of another group, refers to an alkyl or loweralkyl group, as defined herein (and thus including substituted versions such as polyalkoxy), appended to the parent molecular moiety through an oxy group, —O—. Representative examples of alkoxy include, but are not limited to, methoxy, ethoxy, propoxy, 2-propoxy, butoxy, tert-butoxy, pentyloxy, hexyloxy and the like.

“Halo” as used herein refers to any suitable halogen, including —F, —Cl, —Br, and —I.

“Mercapto” as used herein refers to an —SH group.

“Azido” as used herein refers to an —N₃ group.

“Cyano” as used herein refers to a —CN group.

“Formyl” as used herein refers to a —C(O)H group.

“Carboxylic acid” as used herein refers to a —C(O)OH group.

“Hydroxyl” as used herein refers to an —OH group.

“Nitro” as used herein refers to an —NO₂ group.

“Acyl” as used herein alone or as part of another group refers to a —C(O)R radical, where R is any suitable substituent such as aryl, alkyl, alkenyl, alkynyl, cycloalkyl or other suitable substituent as described herein.

“Alkylthio” as used herein alone or as part of another group, refers to an alkyl group, as defined herein, appended to the parent molecular moiety through a thio moiety, as defined herein. Representative examples of alkylthio include, but are not limited, methylthio, ethylthio, tert-butylthio, hexylthio, and the like.

“Amino” as used herein means the radical —NH₂.

“Alkylamino” as used herein alone or as part of another group means the radical —NHR, where R is an alkyl group.

“Arylalkylamino” as used herein alone or as part of another group means the radical —NHR, where R is an arylalkyl group.

“Disubstituted-amino” as used herein alone or as part of another group means the radical —NR_(a)R_(b), where R_(a) and R_(b) are independently selected from the groups alkyl, haloalkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, heterocyclo, heterocycloalkyl.

“Acylamino” as used herein alone or as part of another group means the radical —NR_(a)R_(b), where R_(a) is an acyl group as defined herein and R_(b) is selected from the groups hydrogen, alkyl, haloalkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, heterocyclo, heterocycloalkyl.

“Acyloxy” as used herein alone or as part of another group means the radical —OR, where R is an acyl group as defined herein.

“Ester” as used herein alone or as part of another group refers to a —C(O)OR radical, where R is any suitable substituent such as alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

“Amide” as used herein alone or as part of another group refers to a —C(O)NR_(a)R_(b) radical, where R_(a) and R_(b) are any suitable substituent such as alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

“Sulfoxyl” as used herein refers to a compound of the formula —S(O)R, where R is any suitable substituent such as alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

“Sulfonyl” as used herein refers to a compound of the formula —S(OXO)R, where R is any suitable substituent such as alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

“Sulfonate” as used herein refers to a compound of the formula —S(OXO)OR, where R is any suitable substituent such as alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

“Sulfonic acid” as used herein refers to a compound of the formula —S(OXO)OH.

“Sulfonamide” as used herein alone or as part of another group refers to a —S(O)₂NR_(a)R_(b) radical, where R_(a) and R_(b) are any suitable substituent such as H, alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

“Urea” as used herein alone or as part of another group refers to an —N(R_(c))C(O)NR_(a)R_(b) radical, where R_(a), R_(b) and R_(c) are any suitable substituent such as H, alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

“Alkoxyacylamino” as used herein alone or as part of another group refers to an —N(R_(a))C(O)OR_(b) radical, where R_(a), R_(b) are any suitable substituent such as H, alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

“Aminoacyloxy” as used herein alone or as part of another group refers to an —OC(O)NR_(a)R_(b) radical, where R_(a) and R_(b) are any suitable substituent such as H, alkyl, cycloalkyl, alkenyl, alkynyl or aryl.

1. Active Compounds/Active Agents.

Active compounds or active agents of the present invention are, in general, porphyrin compounds. The active compounds include superoxide dismutase (SOD) mimetic prophryin compounds, and/or redox active porphyrin compounds.

Examples of porphyrin active compounds, and methods of making the same, include but are not limited to those set forth in U.S. Pat. No. 8,470,808 to Piganelli et al.; U.S. Pat. No. 8,183,364 to Batinic-Haberle et al., U.S. Pat. No. 6,916,799 to Fridovich et al.; U.S. Pat. No. 6,479,477 to Crapo et al.; U.S. Pat. No. 6,583,132 to Crapo et al. and in US Patent Application Pub. No. US 2012/0065181 to Warner et al.; the disclosures of which are incorporated by reference herein in their entirety.

Examples of active compounds include but are not limited to compounds of Formula I:

wherein:

each R is independently substituted or unsubstituted aryl, heteroaryl, cycloalkyl, or heterocycloalkyl;

each A is an independently selected hydrogen, or an electron-withdrawing or electron donating group (e.g., e.g., is halogen, NO₂ or CHO),

M is a metal, e.g., selected from the group consisting of manganese, iron, copper, cobalt, nickel and zinc, or is absent (in which case a hydrogen is added to each of the two nitrogens required to correct valency), and

Z⁻ is a counterion.

In some embodiments of Formula I above, each R is preferably heteroaryl or heterocycloalkyl, particularly those containing at least one or two nitrogen atoms in the heterocyclic ring (e.g., pyrrolyl, imidazolyl, triazolyl, pyridyl, pyrimidyl, triazinyl, oxazolyl, thiazolyl, oxazinyl, thiazinyl, oxathiazinyl, etc.), in some embodiments wherein at least one of which nitrogen atoms (or in some embodiments at least two of which nitrogen atoms) are optionally but preferably substituted (e.g., quaternized) with a substituent such as described in connection with heterocyclic groups above (e.g., substituted with alkyl, alkoxyalkyl, etc.).

Still more particular examples of the foregoing active compounds include but are not limited to those set forth below.

A. Alkyl Substituted Imidazole Porphyrins.

In some embodiments the active compound has a structure of Formula A1 or A2:

wherein:

each R is C₁₋₁₂ alkyl (straight chain or branched), more preferably C₂₋₆ alkyl, and most preferably ethyl, propyl, butyl, or pentyl (straight chain or branched);

each A is, independently, hydrogen or an electron withdrawing group (e.g., halogen, NO₂ or CHO),

M is metal selected from the group consisting of manganese, iron, copper, cobalt, nickel and zinc, and

Z⁻ is a counterion.

In some embodiments the active compound has the formula:

wherein Z⁻ is a counterion.

B. Alkyl Substituted Pyridyl Porphyrins

In some embodiments the active compound has a structure of Formula B1 or B2:

wherein:

each R is C₁₋₁₂ alkyl (straight chain or branched), more preferably C₂₋₆ alkyl, and most preferably ethyl, propyl, butyl, or pentyl (straight chain or branched);

each A is, independently, hydrogen or an electron withdrawing group (e.g., halogen, NO₂ or CHO),

M is metal selected from the group consisting of manganese, iron, copper, cobalt, nickel and zinc, and

Z⁻ is a counterion.

In some embodiments the compound has a structure of the Formula V:

wherein each R, A, M and Z is as given in connection with Formula B1 and B2 above.

In some embodiments the compound has the structure:

wherein Z⁻ is a counterion.

C. Alkoxyalkyl Substituted Pyridyl Porphyrins.

In some embodiments the active compound has a structure of Formula C1 or C2:

wherein:

each R is —(CH₂)_(m)CH₂OX;

m is 1 or 2, preferably 1;

X is C₁₋₁₂ alkyl (straight chain or branched), more preferably C₂₋₆ alkyl, and most preferably ethyl, propyl, butyl, or pentyl (straight chain or branched).

each A is, independently, hydrogen or an electron withdrawing group (e.g., halogen, NO₂ or CHO),

M is metal selected from the group consisting of manganese, iron, copper, cobalt, nickel and zinc, and

Z⁻ is a counterion.

In some embodiments the compound has a structure of the Formula V:

wherein each R, A, M and Z is as given in connection with Formula C1 and C2 above.

In some embodiments the compound has the structure:

wherein Z⁻ is a counterion.

D. Salts.

The active compounds disclosed herein can, as noted above, be prepared in the form of their salts or pharmaceutically acceptable salts, e.g., to provide a compound or composition including a counterion as noted above. Pharmaceutically acceptable salts are salts that retain the desired biological activity of the parent compound and do not impart undesired toxicological effects. Examples of such salts are (a) acid addition salts formed with inorganic acids, for example hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid and the like; and salts formed with organic acids such as, for example, acetic acid, oxalic acid, tartaric acid, succinic acid, maleic acid, fumaric acid, gluconic acid, citric acid, malic acid, ascorbic acid, benzoic acid, tannic acid, palmitic acid, alginic acid, polyglutamic acid, naphthalenesulfonic acid, methanesulfonic acid, p-toluenesulfonic acid, naphthalenedisulfonic acid, polygalacturonic acid, and the like; (b) salts formed from elemental anions such as chlorine, bromine, and iodine, and (c) salts derived from bases, such as ammonium salts, alkali metal salts such as those of sodium and potassium, alkaline earth metal salts such as those of calcium and magnesium, and salts with organic bases such as dicyclohexylamine and N-methyl-D-glucamine.

2. Pharmaceutical Formulations.

The active compounds described above may be formulated for administration in a pharmaceutical carrier in accordance with known techniques. See, e.g., Remington, The Science And Practice of Pharmacy (9^(th) Ed. 1995). In the manufacture of a pharmaceutical formulation according to the invention, the active compound (including the physiologically acceptable salts thereof) is typically admixed with, inter alia, an acceptable carrier. The carrier must, of course, be acceptable in the sense of being compatible with any other ingredients in the formulation and must not be deleterious to the patient. The carrier may be a solid or a liquid, or both, and is preferably formulated with the compound as a unit-dose formulation, for example, a tablet, which may contain from 0.01 or 0.5% to 95% or 99% by weight of the active compound. One or more active compounds may be incorporated in the formulations of the invention, which may be prepared by any of the well known techniques of pharmacy comprising admixing the components, optionally including one or more accessory ingredients.

The formulations of the invention include those suitable for oral, rectal, topical, buccal (e.g., sub-lingual), vaginal, parenteral (e.g., subcutaneous, intramuscular, intradermal, or intravenous), topical (i.e., both skin and mucosal surfaces, including airway surfaces) and transdermal administration, although the most suitable route in any given case will depend on the nature and severity of the condition being treated and on the nature of the particular active compound which is being used.

Formulations suitable for oral administration may be presented in discrete units, such as capsules, cachets, lozenges, or tablets, each containing a predetermined amount of the active compound; as a powder or granules; as a solution or a suspension in an aqueous or non-aqueous liquid; or as an oil-in-water or water-in-oil emulsion. Such formulations may be prepared by any suitable method of pharmacy which includes the step of bringing into association the active compound and a suitable carrier (which may contain one or more accessory ingredients as noted above). In general, the formulations of the invention are prepared by uniformly and intimately admixing the active compound with a liquid or finely divided solid carrier, or both, and then, if necessary, shaping the resulting mixture. For example, a tablet may be prepared by compressing or molding a powder or granules containing the active compound, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing, in a suitable machine, the compound in a free-flowing form, such as a powder or granules optionally mixed with a binder, lubricant, inert diluent, and/or surface active/dispersing agent(s). Molded tablets may be made by molding, in a suitable machine, the powdered compound moistened with an inert liquid binder.

Formulations suitable for buccal (sub-lingual) administration include lozenges comprising the active compound in a flavoured base, usually sucrose and acacia or tragacanth; and pastilles comprising the compound in an inert base such as gelatin and glycerin or sucrose and acacia.

Formulations of the present invention suitable for parenteral administration comprise sterile aqueous and non-aqueous injection solutions of the active compound(s), which preparations are preferably isotonic with the blood of the intended recipient. These preparations may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient. Aqueous and non-aqueous sterile suspensions may include suspending agents and thickening agents. The formulations may be presented in unit\dose or multi-dose containers, for example sealed ampoules and vials, and may be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example, saline or water-for-injection immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described. For example, in one aspect of the present invention, there is provided an injectable, stable, sterile composition comprising an active compound(s), or a salt thereof, in a unit dosage form in a sealed container. The compound or salt is provided in the form of a lyophilizate which is capable of being reconstituted with a suitable pharmaceutically acceptable carrier to form a liquid composition suitable for injection thereof into a subject. The unit dosage form typically comprises from about 10 mg to about 10 grams of the compound or salt. When the compound or salt is substantially water-insoluble, a sufficient amount of emulsifying agent which is physiologically acceptable may be employed in sufficient quantity to emulsify the compound or salt in an aqueous carrier. One such useful emulsifying agent is phosphatidyl choline.

Formulations suitable for rectal administration are preferably presented as unit dose suppositories. These may be prepared by admixing the active compound with one or more conventional solid carriers, for example, cocoa butter, and then shaping the resulting mixture.

Formulations suitable for topical application to the skin preferably take the form of an ointment, cream, lotion, paste, gel, spray, aerosol, or oil. Carriers which may be used include petroleum jelly, lanoline, polyethylene glycols, alcohols, transdermal enhancers, and combinations of two or more thereof.

Formulations suitable for transdermal administration may be presented as discrete patches adapted to remain in intimate contact with the epidermis of the recipient for a prolonged period of time. Formulations suitable for transdermal administration may also be delivered by iontophoresis (see, for example, Pharmaceutical Research 3 (6):318 (1986)) and typically take the form of an optionally buffered aqueous solution of the active compound. Suitable formulations comprise citrate or bis\ris buffer (pH 6) or ethanol/water and contain from 0.1 to 0.2M active ingredient.

Further, the present invention provides liposomal formulations of the compounds disclosed herein and salts thereof. The technology for forming liposomal suspensions is well known in the art. When the compound or salt thereof is an aqueous-soluble salt, using conventional liposome technology, the same may be incorporated into lipid vesicles. In such an instance, due to the water solubility of the compound or salt, the compound or salt will be substantially entrained within the hydrophilic center or core of the liposomes. The lipid layer employed may be of any conventional composition and may either contain cholesterol or may be cholesterol-free. When the compound or salt of interest is water-insoluble, again employing conventional liposome formation technology, the salt may be substantially entrained within the hydrophobic lipid bilayer which forms the structure of the liposome. In either instance, the liposomes which are produced may be reduced in size, as through the use of standard sonication and homogenization techniques.

Of course, the liposomal formulations containing the compounds disclosed herein or salts thereof, may be lyophilized to produce a lyophilizate which may be reconstituted with a pharmaceutically acceptable carrier, such as water, to regenerate a liposomal suspension.

Other pharmaceutical compositions may be prepared from the water-insoluble compounds disclosed herein, or salts thereof, such as aqueous base emulsions. In such an instance, the composition will contain a sufficient amount of pharmaceutically acceptable emulsifying agent to emulsify the desired amount of the compound or salt thereof. Particularly useful emulsifying agents include phosphatidyl cholines, and lecithin.

In addition to active compound(s), the pharmaceutical compositions may contain other additives, such as pH-adjusting additives. In particular, useful pH-adjusting agents include acids, such as hydrochloric acid, bases or buffers, such as sodium lactate, sodium acetate, sodium phosphate, sodium citrate, sodium borate, or sodium gluconate. Further, the compositions may contain microbial preservatives. Useful microbial preservatives include methylparaben, propylparaben, and benzyl alcohol. The microbial preservative is typically employed when the formulation is placed in a vial designed for multidose use. Of course, as indicated, the pharmaceutical compositions of the present invention may be lyophilized using techniques well known in the art.

3. Dosage and Routes of Administration.

As noted above, the present invention provides pharmaceutical formulations comprising the active compounds (including the pharmaceutically acceptable salts thereof), in pharmaceutically acceptable carriers for oral, rectal, topical, buccal, parenteral, intramuscular, intradermal, or intravenous, and transdermal administration.

The effective amount (e.g., therapeutically effective or treatment effective amount) or dosage of any specific active compound as described herein, for use in any specific method as described herein, will vary depending on factors such as the condition being treated, the route of administration, the general condition of the subject (e.g., age, gender, weight, etc.), etc. In general (e.g., for oral or parenteral administration), the dosage may be from about 0.01, 0.05, or 0.1 milligram per kilogram subject body weight (mg/kg), up to about 1, 5, or 10 mg/kg. For topical administration, the active agent may be included in a pharmaceutically acceptable composition to be applied in any suitable amount, typically from 0.01, 0.1, or 1 percent by weight, up to 10, 20, or 40 percent by weight, or more, of the weight of the composition, again depending on factors such as the condition being treated, condition of the subject, etc.

The active agents described herein may be administered directly or through the administration to the subject of a pharmaceutically acceptable prodrug which is in turn converted to the active agent in vivo. The term “prodrug” refers to compounds that are rapidly transformed in vivo to yield the parent compound of the above formulae, for example, by hydrolysis in blood. A thorough discussion is provided in T. Higuchi and V. Stella, Prodrugs as Novel delivery Systems, Vol. 14 of the A.C.S. Symposium Series and in Edward B. Roche, ed., Bioreversible Carriers in Drug Design, American Pharmaceutical Association and Pergamon Press, 1987, both of which are incorporated by reference herein. See also U.S. Pat. No. 6,680,299 Examples include a prodrug that is metabolized in vivo by a subject to an active drug having an activity of active compounds as described herein, wherein the prodrug is an ester of an alcohol or carboxylic acid group, if such a group is present in the compound; an acetal or ketal of an alcohol group, if such a group is present in the compound; an N-Mannich base or an imine of an amine group, if such a group is present in the compound; or a Schiff base, oxime, acetal, enol ester, oxazolidine, or thiazolidine of a carbonyl group, if such a group is present in the compound, such as described in U.S. Pat. No. 6,680,324 and U.S. Pat. No. 6,680,322.

4. Biofilm Treatment.

Active agents as described herein can be used to treat, control, inhibit or disrupt biofilm formation in vivo or in vitro in like manner as described in connection with other biofilm dispersing or disrupting agents, including but not limited to those described in U.S. Pat. No. 8,513,305 to Davies, the disclosure of which is incorporated herein by reference in its entirety.

Biofilms formed from any film-forming microorganism (e.g., fungal or bacterial, including gram negative and gram positive bacteria) can be treated by the methods of the present invention. Examples of such film-forming microorganisms include, but are not limited to, Staphylococcus aureus (e.g., methicillin-resistant Staphylococcus aureus (MRSA)), Haemophilus influenza, Pseudomonas aeruginosa, Burkholdereia cepacla, Escherichia coli, Actinomyces israelii, etc.

Thus an aspect of the present invention relates to a method of treating or preventing a condition mediated by a biofilm in a subject. The method comprises providing a subject having, or susceptible to, a condition mediated by a biofilm produced by a microorganism, whereby the biofilm comprises a matrix and the micro-organism on a surface. Administered to the subject is an active agent as described herein, under conditions effective for the active agent to selectively act on the microorganism and have a suitable biological response without a required direct effect on the matrix. As a result, the condition mediated by a biofilm in the subject is treated, inhibited or prevented. The method of dispersing the biofilm may further include administering to the biofilm, in conjunction with administering the active agent, an antimicrobial treatment. The treatment can be the administration of biocides (e.g., hydrogen peroxide), surfactants, antibiotics, antiseptics, detergents, chelating agents, virulence factor inhibitors, gels, polymers, pastes, edible products, chewable products, ultrasonic treatment, radiation treatment, thermal treatment, and/or mechanical treatment.

An additional aspect of the present invention relates to a method of treating or inhibiting formation of a biofilm on a surface. This method involves providing a surface having or being susceptible to formation of a biofilm produced by a microorganism, where the biofilm comprises a matrix and the micro-organism on the surface. Administered to the surface is a active agent as described herein under conditions effective for the formation of the biofilm on the surface is treated or inhibited.

In one embodiment, the surface to be treated includes indwelling medical devices, such as catheters (e.g., urinary tract catheters), respirators, ventilators, and intrauterine devices (IUDs). In addition, the surface can be in implanted medical devices, such as stents, artificial valves, joints, sutures, staples, pacemakers, bone implants, pins, vascular grafts, mechanical heart valves, arteriovenous shunts, cerebral spinal fluid shunts, endovascular catheters, penile prostheses, surgical mesh, and orthopedic fixation devices (e.g., plates, rods, screws, etc.). The active agent of the present invention can also be included in surgical glue.

In another embodiment, the surface to be treated includes drains, tubs, kitchen appliances, countertops, shower curtains, grout, toilets, industrial food and beverage production facilities, flooring, and food processing equipment.

In a further embodiment, the surface to be treated is a heat exchanger surface or a filter surface. Thus, treatment provides a means for reducing the degree of biofouling of the heat exchanger or filter.

In a further embodiment, the surface to be treated is a marine structure which includes boats, piers, oil platforms, water intake ports, sieves, and viewing ports. The surface can alternatively be associated with a system for water treatment and/or distribution (e.g., a system for drinking water treatment and/or distributing, a system for pool and spa water treatment, a system for treatment and/or distribution of water in manufacturing operations, and a system for dental water treatment and/or distribution). The surface can also be associated with a system for petroleum drilling, storage, separation, refining and/or distribution (e.g., a petroleum separation train, a petroleum container, petroleum distributing pipes, and petroleum drilling equipment). The active agent can also be included in formulations directed at reducing or eliminating biofilm deposits or biofouling in porous medium, such as with oil and gas bearing geological formations. The treatment may be accomplished by applying a coating containing the active agent, such as paint, to the surface.

The method of inhibiting formation of a biofilm on a surface may further involve administering to the surface, in conjunction with administering the active agent, an antimicrobial treatment. The treatment can be administration of biocides, surfactants, antibiotics, antiseptics, disinfectants, medicines, detergents, chelating agents, virulence factor inhibitors, ultrasonic treatment, radiation treatment, thermal treatment, and mechanical treatment. In one embodiment, the active agent and the antimicrobial treatment are administered simultaneously. In another embodiment, the active agent and antimicrobial treatment are administered separately.

The active agent can be impregnated in a surface in order to inhibit formation of a biofilm on the surface. Alternatively, the active agent can be in a copolymer or a gel coating over the surface.

The present invention also relates to a method of treating subjects with burns. The method involves administering the active agent according to the present invention, under conditions effective to treat burns in the subject. A specific application of the invention provides a topical dressing for burn patients comprising dispersion inducing molecules or their natural or synthetic analogs to prevent the development of infectious biofilms or to disperse the cells of existing infectious biofilms.

The present invention further relates to a method of treating and/or preventing dental plaque, dental carries, gingival disease, periodontal disease, and oral infection in a subject. The method involves treating the oral cavity of the subject with the active agent according to the present invention. Treating can be carried out with a dentifrice, mouthwash, dental floss, gum, strip, toothpaste, a toothbrush containing the active agent, and other preparations containing the active agent. The composition may also contain other compounds known in the dental arts that are typically added to dental compositions. For example, the active agent composition may also include fluoride, desensitizing agents, anti-tartar agents, anti-bacterial agents, remineralization agents, whitening agents, and anti-caries agents.

The amount of active agent present will vary dependent on the dental composition that contains the active agent. It has been found that the active agent is active over a wide range of concentrations against oral bacteria. For instance, the active agent may be present in an amount ranging from 0.1 nM to 10 mM. However, lower and higher concentrations may be used depending on the dental composition, the other components present in the active agent composition, and various other factors appreciated by those of skill in the art. The known properties of the active agent, such as its fatty acid characteristics and its hydrophobicity, will assist a skilled artisan in determining how much of the active agent should be used, determining how the compound will chemically interact with other components, and providing other useful information about the compound.

Specific dental applications and dental compositions are contemplated in this invention. In this regard, the invention relates to a toothbrush containing a active agent composition. Toothbrushes, as is well known in the art, contain a plurality of bristles and a solid support on which the bristles are mounted, where the solid support includes a brush head having a plurality of tuft holes that receive the bristles. Variations and modifications of the basic toothbrush are well known in the art. See, for example, U.S. Pat. No. 7,251,849, herein incorporated by reference in its entirety.

The active agent of this invention has a chemical formula as set forth above. Additional components that may be included in the active agent compositions are also set forth above. The active agent composition may be incorporated in the various parts of the toothbrush by means known in the art. For instance, the active agent composition may be contained in the tuft holes of the toothbrush. See U.S. Pat. No. 5,141,290, herein incorporated by reference in its entirety, for an example of how a composition can be contained within the tuft holes of a toothbrush. Alternatively, the active agent composition may be coated or embedded in the bristles of the toothbrush.

Other parts of the toothbrush may also be coating or embedded with the active agent composition, including any parts of the toothbrush that supplement the bristles and are designed to be contacted with the oral cavity. For example, it is common for toothbrushes to contain rubber paddles, tongue cleaners, or other pieces extended from the head for the purposes of being contacted with the tooth, tongue, gums, or other areas of the oral cavity. These parts may be embedded with the active agent composition and, optionally, a surfactant, biocide, and/or other additive discussed above.

To assist in controlling the release of the active agent from the toothbrush, the active agent composition may contain an agent that interacts with the active agent to assist in the controlled release. The agent may interact with the active agent in a manner that the release is either accelerated or prolonged, depending on the desired use. The level of controlled release can also depend on how easily or difficult the active agent adheres to the portion of the toothbrush that it is applied to. In a preferred embodiment, the active agent is slowly released from the toothbrush over repeated brushings. Agents that enable the slow release of an active ingredient are well known to those of skill in the art.

The controlled release may also be effectuated by encapsulating the active agent in an encapsulated system that allows a controlled release. In this embodiment, the active agent composition is preferably in the form of a plurality of small microspheres that encapsulate the active agent. The microspheres can have an outer coating of dissolvable material that enables the active agent to slowly release over repeated brushings. Suitable microspheres include those disclosed in U.S. Pat. No. 5,061,106, herein incorporated by reference in its entirety.

This invention also relates to a toothpaste composition that contains (a) fluoride and/or a remineralization agent; (b) an orally-accepted vehicle; and (c) a active agent composition. The active agent of this invention has a chemical formula as set forth above. Additional components that may be included in the active agent compositions are also set forth above. Often, toothpastes also contain sodium lauryl sulfate or other sulfates.

Fluoride in its various forms is a common active ingredient in toothpaste to prevent cavities and promote the formation of dental enamel and bones. Any fluoride source, such as fluoride salts may be used in the toothpaste of this invention. Preferably, the fluoride is sodium fluoride (NaF) or sodium monofluorophosphate. Typically, the amount of fluoride present in the toothpaste ranges from 100 to 5000 parts per million fluoride ion, preferably 1000 to 1100 parts per million.

In certain instances, it is preferable to replace or supplement the fluoride with a remineralization agent. Remineralization, in the context of dental usage, generally refers to treating the teeth so as to prevent dental caries, or decrease their chance of occurring, and otherwise enhance the teeth so that they can return to their original, healthy state. While fluoride can be considered a remineralization agent, other agents often take the place of fluoride or supplement fluoride to provide the toothpaste with a stronger cleansing or remineralization properties. Common remineralization agents are calcium salts, such as calcium phosphate, calcium sulfate, anhydrous calcium sulfate, calcium sulfate hemihydrate, calcium sulfate dihydrate, calcium malate, calcium tartrate, calcium malonate, and calcium succinate. Hydroxyapitate nanocrystals and zinc compounds have also been shown to be effective remineralization agents.

The orally-accepted vehicle may be any vehicle known in the art that can be used to deliver the fluoride and/or remineralization agent, and active agent to the teeth of a patient. The orally-accepted vehicle may also be glycerin, propylene glycol, polyethylene glycol, triglyceride, diglyceride, mineral oil, organic oils, essential oils, fatty vegetable oils, and combinations thereof. Often these vehicles are used in combination with water or a water-based solvent.

The toothpaste composition may contain other components of toothpastes well known in the art. For instance, the toothpaste composition may contain baking soda, enzymes, vitamins, herbs, calcium compounds such as calcium sodium phosphosilicate, coloring agents, and/or flavoring agents. Desensitizing agents may also be added. As known in the art, desensitizing agents can reduce sensitivity in teeth by treating sensitivities caused by demineralization or suppressing the sensitivity symptoms by desensitizing the nerves. The composition may also contain an antibacterial or an antiplaque agent. Antibacterial agents are preferable included in the composition to prevent gingivitis, periodontitis, and other oral diseases. Suitable antibacterial agents include triclosan, zinc chloride, chlorhexidine, benzethonium chloride, and cetyl pyridinium chloride.

This invention also relates to an oral composition for treating and/or preventing dental plaque, gingival diseases, periodontal diseases, and/or oral infection. The oral composition contains an orally-accepted vehicle and a active agent composition. The active agent of this invention has a chemical formula as set forth above. Additional components that may be included in the active agent compositions are also set forth above.

The oral composition can be various compositions in the field of dental hygiene known to those in the art. For instance, the oral composition may be a mouthwash, breath spray, dentifrice, tooth powder, whitening strips, or prophylaxis paste. As is well known in the art, mouthwashes are commonly used to help remove mucous and food particles in the oral cavity or throat. Mouthwashes typically contain antiseptic and/or anti-plaque components to kill the bacterial plaque that causes caries, gingivitis, and bad breath. They can also contain anti-cavity components, such as fluoride, to protect against tooth decay. Suitable mouthwash components may be found in U.S. Pat. No. 5,968,480, herein incorporated by reference in its entirety.

Likewise, the same or similar antiseptic, anti-plaque, and anti-cavity components can be used in breath sprays, dentifrices, including gel dentifrices, tooth powders, whitening strips, and prophylaxis pastes. Suitable breath spray compositions are disclosed in U.S. Pat. No. 7,297,327; suitable tooth powder compositions, such as those used in tooth bleaching compositions, are disclosed in U.S. Pat. No. 5,989,526; suitable whitening strips are disclosed in U.S. Pat. No. 6,514,483; and suitable dentifrices and prophylaxis paste compositions, including dental abrasives, are disclosed in U.S. Pat. No. 5,939,051, all of which are herein incorporated by reference in their entirety.

The ingredients of orally-accepted vehicle are similar to those discussed above. However, the orally-accepted vehicle will vary depending on the desired consistency and desired end product of the oral composition. For instance, a mouthwash is in a liquid form, so liquid carriers, typically carriers having a high percentage of water, should be used. On the other hand, a gel dentifrice should be in the form of a gel and would utilize gelling agents or other carriers that enable the final product to be in the form of a gel. The orally-accepted vehicle should have properties that both allow the active agent composition to be delivered while also providing the final product with the desired consistency.

The oral composition may also be in the form of chewing gum, a breath strip, a lozenge, or a breath mint. Chewing gum is typically a combination of a water-insoluble phase, or gum base, and a water-soluble phase of sweeteners, flavoring and/or food coloring. Other components may also be added to the gum, including breath-freshening additives such as zinc and phosphate salts, teeth-whitening additives such as silica, and plaque-reducing additives to moderate dental plaque. Suitable gum compositions may be found in U.S. Pat. Nos. 6,416,744 and 6,592,849, both of which are herein incorporated by reference in their entirety.

Breath strips are similar to chewing gum, except that the strips are designed to dissolve in the mouth, often absorbed through the tongue. The strips can deliver bioactive ingredients to freshen the mouth as well functional bioactive ingredients, such as vitamins, minerals, supplements, pharmaceuticals, and vaccines.

Lozenges and breath mints are typically discoid-shaped solids that contain a therapeutic agent in a flavored base. The base may be a hard sugar candy, glycerinated gelatin or combination of sugar with sufficient mucilage to give the composition requisite form. The active agent may represent the therapeutic agent, or it may be added in addition to therapeutic agents known in the art. Suitable lozenge and breath mint compositions are disclosed in U.S. Pat. No. 7,025,950, herein incorporated by reference in its entirety.

The oral composition may also be in the form of a cleaning preparation for a dental apparatus that is placed in the oral cavity. Dental apparatuses such as dentures, dental dams, and certain types of orthodontic braces are placed in the oral cavity for a period of time, and then periodically removed for cleaning. The cleaning composition used to clean the dental apparatuses should function in its customary manner of cleaning the apparatus, but may also contain therapeutic agents that can assist in treating or preventing dental plaque, gingival diseases, periodontal diseases, and oral infection when the dental apparatuses are in contact with the oral cavity. Cleaning compositions such as effervescent cleansers made with alkaline mixtures containing a chlorine compounds and the like are known in the art. Suitable cleaning compositions for dental apparatuses are disclosed in U.S. Pat. No. 3,936,385, herein incorporated by reference in its entirety. The active agent may be added to the cleaning compositions in a manner than enables it to coat the dental apparatus upon contact. After the dental apparatus has been introduced into the oral cavity, the active agent can interact with the teeth and other elements of the oral cavity in a therapeutically effective manner, i.e. to prevent dental plaque, gingival diseases, periodontal diseases, and/or oral infection.

This invention also relates to an article for oral use comprising a dental article and a active agent. The active agent has the chemical formula set forth above, and is coated on, encapsulated in, or impregnated in the dental article. Additional components that may be included in the active agent compositions are also set forth above.

Various dental articles known in the art may used in this embodiment of the invention. In one embodiment, the dental article is a dental floss. Any fiber known in the art may be used in the dental floss. Suitable fibers include polyamides (such as nylon), polyesters, polypropylenes, polytetrafluoroethylenes, cellulose, and cotton. Nylon and polytetrafluoroethylene fibers are the most common fibers used in dental floss and represent preferred fibers. Suitable dental flosses are disclosed in U.S. Pat. Nos. 6,270,890 and 6,289,904, both of which are herein incorporated by reference in their entirety. The active agent composition may be impregnated into the fiber, coated on the fiber, or otherwise incorporated into the dental floss.

The dental floss may be coated or impregnated with a wax or other hydrophobic substance for ease of use during the flossing process. Suitable waxes include microcrystalline waxes, beeswax, paraffin waxes, carnauba waxes, and polyethylene waxes. The active agent composition may be coated onto the dental floss as part of the wax layer, as a second or additional layer in conjunction with the wax layer, or applied to the fiber as discussed above.

The dental article may be a toothpick that is impregnated with or coated with the active agent composition. Toothpicks may be made from natural products, such as wood, or artificial components, including various plastics. Suitable toothpicks are disclosed in U.S. Pat. No. 7,264,005, herein incorporated by reference in its entirety.

The dental article may also be a dental appliance such as a dental aspirator, bite block, dental dam, tongue stabilizer, tongue deflector, or any other piece of dental equipment that a dentist or dental assistant may use in the mouth of a patient. A discussion of dental appliances may be found in U.S. Pat. Nos. 4,865,545 and 5,152,686, both of which are herein incorporated by reference. The portion of the dental appliance that comes into contact with the oral cavity of a patient may be coated with the active agent composition.

The dental article may also be a dental construct, such as a veneers, crowns, inlays, onlays, or bridges that are placed on the teeth. Dental constructs are typically made of metal alloys, porcelain, ceramic, amalgam, acrylate polymers, or a combination of these materials. Suitable dental constructs are disclosed in U.S. Pat. No. 7,229,286, herein incorporated by reference in its entirety. The active agent composition may be embedded in the composition used to make the dental construct. Alternatively, the active agent composition may be coated on the dental construct after it has been prepared.

This invention also relates to an aqueous composition applied to the oral cavity with the use of a dental article, comprising water and a active agent composition. Various dental articles are attached to or designed to be used with a water line so that water can be distributed through the dental article, and then routed from the dental article to the oral cavity of a subject. Suitable dental articles include dental water lines, dental water picks, and the like.

While tap water or purified water may be used in these type of dental devices, the water source may also be supplemented with additives so that the water delivers the additives to the oral cavity of the subject when used with the dental article. In this case, the additive supplemented to the water is a active agent composition.

Dental water lines and dental water picks are known in the art and commonly used by dentists and dental assistants. A discussion of different types of dental water lines and their different applications may be found in U.S. Pat. No. 5,785,523, herein incorporated by reference in its entirety. Suitable water picks are disclosed in U.S. Pat. No. 4,257,433, herein incorporated by reference in its entirety.

The present invention also relates to a method of cleaning and/or disinfecting contact lenses. The method involves treating contact lenses with a cleaning and/or disinfecting solution containing the active agent according to the present invention. The contact lens may be treated in this manner while being stored in solution or while being used in vivo. Alternatively, the active agent can be used in eye drops.

The present invention further relates to a method of treating and/or preventing acne or other biofilm-associated skin infections on the skin of a subject. The method involves treating the skin of the subject with the active agent according to the present invention under conditions effective to treat and/or prevent the acne or biofilm-associated skin infections. The active agent may be present in an ointment, cream, liniment, salves, shaving lotion, or aftershave. It may also be present in a powder, cosmetic, ointment, cream, liquid, soap, gel, cosmetic applicator, and/or solid, woven or non-woven material intended to contact or be proximate with the skin.

The present invention also relates to a method of treating and/or preventing a chronic biofilm-associated disease in a subject. The method involves administering to the subject the active agent according to the present invention under conditions effective to treat and/or prevent the chronic biofilm-associated disease. The chronic biofilm-associated diseases to be treated and/or prevented include, but are not limited to, middle ear infections, osteomyelitis, prostatitis, colitis, vaginitis, urethritis, arterial plaques, sinovial infections, infections along tissue fascia, respiratory tract infections (e.g., infections associated with lung infections of cystic fibrosis patients, pneumonia, pleurisy, pericardial infections), genito-urinary infections, and gastric or duodenal ulcer infections. For gastric or duodenal ulcers caused by Helicobacter pylori, the active agent will need to function at a pH of below 5.5. The active agent may be administered in combination with an antimicrobial agent, such as biocides, surfactants, antibiotics, antiseptics, detergents, chelating agents, or virulence factor inhibitors. In the case of gastric therapies, acid reducing therapies, such as antacids, proton pump inhibitors, antihistamines, and the like may also be employed.

The present invention also relates to a composition comprising one or more active agents and one or more additive components. These additive components are selected from the group consisting of biocides, surfactants, antibiotics, antiseptics, detergents, chelating agents, virulence factor inhibitors, gels, polymers, pastes, edible products, and chewable products. The composition is formulated so that when it is contacted with a biofilm produced by a microorganism, where the biofilm comprises a matrix and microorganism on a surface, the active agent selectively acts on the microorganism and has a suitable biological response without a required direct effect to disrupt the matrix.

Another aspect of the present invention relates to a method of treating or preventing a condition mediated by a biofilm in a subject. This method involves providing a subject having, or susceptible to, a condition mediated by a biofilm produced by a microorganism, whereby the biofilm comprises a matrix and the micro-organism on a surface. An active agent is administered to the subject under conditions effective for the active agent to selectively act on the microorganism and have a suitable biological response without a required direct effect on the matrix, whereby the condition mediated by a biofilm in the subject is treated or prevented.

A further embodiment of the present application is directed to a method of treating or inhibiting formation of a biofilm on a surface. This involves providing a surface having or being susceptible to formation of a biofilm produced by a microorganism, whereby the biofilm comprises a matrix and the micro-organism on the surface. An active agent is administered to the surface under conditions effective for the active agent to selectively act on the microorganism and have a suitable biological response without a required direct effect on the matrix, whereby formation of the biofilm on the surface is treated or inhibited.

When the active agents described herein are used in combination with an antibiotic, any suitable antibiotic may be used (alone, or in combination with other antibiotics). Examples include, but are not limited to, at least one of aminoglycoside; amikacin; gentamicin; kanamycin; neomycin; netilmicin; steptomycin; tobramycin; ansamycin; geldanamycin; herbimycin; carbacephem; loracarbef; carbacepenem; ertapenem; doripenem; imipenem/cilastatin; meropenem; cephalosporin; cefadroxil; cefazolin; cefalotin or cefalothin; cefalexin; cefaclor; cefamandole; cefoxitin; cefprozil; cefuroxime; cefixime; cefdinir; cefditoren; cefoperazone; cefotaxime; cefpodoxime; ceftazidime; ceftibuten; ceftizoxime; ceftriaxone; cefepime; ceftobiprole; glycopeptide; teicoplanin; vancomycin; macrolide; azithromycin; clarithromycin; dirithromycin; erythromicin; roxithromycin; troleandomycin; telithromycin; spectinomycin; monobactam; aztreonam; penicillin; amoxicillin; ampicillin; azlocillin; carbenicillin; cloxacillin; dicloxacillin; flucloxacillin; mezlocillin; meticillin; nafcillin; oxacillin; penicillin, piperacillin, ticarcillin; bacitracin; colistin; polymyxin B; quinolones; ciprofloxacin; enoxacin; gatifloxacin; levofloxacin; lomefloxacin; moxifloxacin; norfloxacin; ofloxacin; trovafloxacin; sulfonamide; mafenide; prontosil (archaic); sulfacetamide; sulfamethizole; sufanilimide (archaic); sulfasalazine; sulfisoxazole; trimethoprim; trimethoprim-sulfamethoxazole (co-trimoxazole) (TMP-SMX); tetracycline; demeclocycline; doxycycline; minocycline; oxytetracycline; tetracycline; arsphenamine; chloramphenicol; clindamycin; lincomycin; ethambutol; fosfomycin; fusidic acid; furazolidone; isoniazid; linezolid; metronidazole; mupirocin; nitrofuantoin; platensimycin; purazinamide; quinupristin/dalfopristin; rifampin or rifampicin; or tinidazole. See, e.g., U.S. Pat. No. 8,211,656.

The present invention is explained in greater detail in the following non-limiting Examples.

Example 1 Inhibition of Staphylococcus Aureus Biofilm Formation In Vitro

Biofilm forming S. aureus was grown in 96 well plates for 24 hours to allow a biofilm to form. Wells were allowed to grow an additional 24 hours after being treated with either PBS or SOD mimetic (Mn (III) tetrakis (N-ethylpyridinium-2-yl) porphyrin, or “MnTE-PyP”). Biofilm intensity was measured using absorbance at 600 nm. Results are shown in FIG. 1. These results showed that MnTE disrupts established S. aureus biofilms in culture. (p<0.0001)

Example 2 Biofilm Disruption In Vivo in a Mouse Infected Fracture Model

A total of 20 C57B5 male mice were randomly divided into four treatment groups after surgery to create an infected femur fracture (midshaft femur fracture fixed with intramedullary 23 g needle and with 10⁴ S. aureus added to the fracture site):

1. Control (no active treatment),

2. Antibiotic (Keflex) starting 24 hours post-surgery,

3. SOD mimetic MnTE-2-PyP (starting 24 hours post-surgery,

4. Antibiotic plus MnTE-2-PyP (starting 24 hours post-surgery).

(Though not felt to be related to the compound, it is noted that one mouse died in the initial post-operative period from the mimetic only group.)

The mice were followed for 2 weeks and received daily access to Keflex in drinking water at 250 m/ml dose for the antibiotic treated groups and MnTE three time per week (loading dose day 1=5.0 mg/kg SQ, followed by 2.5 mg/kg SQ three times weekly). Mice resumed weight-bearing on post operative day 1 and were weighed three times per week and monitored for complications. Mice were harvested after 2 weeks and femurs were homogenized in PBS, supernatants were serially diluted and plated to determine cell counts. Results are given in FIGS. 2-3.

The foregoing is illustrative of the present invention, and is not to be construed as limiting thereof. The invention is defined by the following claims, with equivalents of the claims to be included therein. 

1. A method of treating or preventing a condition caused by a biofilm in a subject, said method comprising: providing a subject having, or susceptible to, a condition mediated by a biofilm produced by a microorganism, where the biofilm comprises a matrix and the microorganism on a surface and administering to the subject an active agent under conditions effective for the condition caused by a biofilm in the subject to be treated or prevented; wherein said active agent is a compound of Formula I:

wherein: each R is independently selected substituted or unsubstituted aryl, heteroaryl, cycloalkyl, or heterocycloalkyl; each A is an independently selected hydrogen, an electron-withdrawing group, or electron donating group; M is a metal; and Z⁻ is a counterion; or a pharmaceutically acceptable salt thereof.
 2. The method of claim 1 further comprising: administering to the subject, in conjunction with said administering the active agent, an antimicrobial treatment selected from the group consisting of one or more of biocides, surfactants, antibiotics, antiseptics, detergents, chelating agents, virulence factor inhibitors, ultrasonic treatment, radiation treatment, thermal treatment, and mechanical treatment.
 3. The method of claim 1, wherein a subject with a bone fracture is treated (e.g., an open or compound bone fracture).
 4. The method of claim 1, wherein a subject with a burn (e.g., second or third degree burns, including chemical and thermal burns to the skin, eye, etc.) is treated, or other acute or chronic wound (e.g., incision, laceration, ulceration such as a bed sore, etc.)
 5. The method of claim 1, wherein a subject with dental plaque, dental caries, gingival disease, and/or oral infection is treated.
 6. The method of claim 5, wherein said administering is carried out with a dentifrice, mouthwash, dental floss, gum, strip, or brush.
 7. The method of claim 1, wherein a subject with acne or other biofilm-associated skin infections on the skin is treated.
 8. The method of claim 1, wherein a subject with a chronic biofilm-associated disease is treated.
 9. The method of claim 8, wherein the chronic biofilm-associated disease is selected from the group consisting of middle ear infection, osteomyelitis, prostatitis, cystic fibrosis, colitis, vaginitis, urethritis, gastric or duodenal ulcer, ventilation-associated pneumonia, endocarditis, necrotizing fasciitis, biliary tract infection, and infectious kidney stones.
 10. The method of claim 1, wherein a subject with a Staphylococcus aureus (e.g., methicillin-resistant Staphylococcus aureus (MRSA)), Haemophllus influenza, Pseudomonas aeruginosa Burkholderea cepacia Escherichla coil, or Actinomyces israelli infection is treated.
 11. A method of treating or inhibiting formation of a biofilm on a surface, said method comprising: providing a surface having or being susceptible to formation of a biofilm produced by a microorganism, where the biofilm comprises a matrix and the microorganism on the surface, and administering to the surface an active agent under conditions effective for formation of the biofilm on the surface to be treated or inhibited; wherein said active agent is a compound of Formula I:

wherein: each R is independently selected substituted or unsubstituted aryl, heteroaryl, cycloalkyl, or heterocycloalkyl; each A is an independently selected hydrogen, an electron-withdrawing group, or electron donating group; M is a metal; and Z⁻ is a counterion; or a pharmaceutically acceptable salt thereof.
 12. The method of claim 11, wherein the surface is a contact lens.
 13. The method of claim 11, wherein the surface is an indwelling medical device selected from the group consisting of catheters (e.g., urinary tract catheters), respirators, ventilators, and intrauterine devices (IUDs).
 14. The method of claim 11, wherein the surface is an implanted medical device selected from the group consisting of stents, artificial valves, joints, sutures, staples, pacemakers, bone implants, pins, vascular grafts, mechanical heart valves, arteriovenous shunts, cerebral spinal fluid shunts, endovascular catheters, penile prostheses, surgical mesh, and orthopedic fixation devices (e.g., plates, rods, screws, etc.).
 15. The method of claim 11, wherein the surface is selected from the group consisting of drains, tubs, kitchen appliances, countertops, shower curtains, grout, toilets, industrial food and beverage production facilities, flooring, and food processing equipment.
 16. The method of claim 11, wherein the surface is a heat exchanger surface or a filter surface.
 17. The method of claim 11, wherein the surface is a marine structure selected from the group consisting of boats, piers, oil platforms, water intake ports, sieves, and viewing ports.
 18. The method of claim 11, wherein the surface is associated with a system for water treatment and/or distribution.
 19. The method of claim 18, wherein the system for water treatment and/or distribution is selected from the group consisting of a system for drinking water treatment and/or distribution, a system for pool and spa water treatment, a system for treatment and/or distribution of water in manufacturing operations, and a system for dental water treatment and/or distribution.
 20. The method of claim 11, wherein the surface is associated with a system for petroleum drilling, storage, separation, refining, distribution, and/or porous medium from which the petroleum is extracted.
 21. The method claim 20, wherein the system for petroleum drilling, storage, separation, and/or distribution is selected from the group consisting of a petroleum separation train, a petroleum container, petroleum distributing pipes, and petroleum drilling equipment.
 22. The method of claim 11 further comprising: administering to the surface, in conjunction with said administering the active agent, at least one antimicrobial treatment selected from the group consisting of biocides, surfactants, antibiotics, antiseptics, detergents, chelating agents, virulence factor inhibitors, ultrasonic treatment, radiation treatment, thermal treatment, and mechanical treatment.
 23. The method of claim 22, wherein the active agent and the antimicrobial treatment are administered simultaneously.
 24. The method of claim 22, wherein the active agent and antimicrobial treatment are administered separately.
 25. The method of claim 22, wherein the active agent is impregnated in the surface.
 26. The method of claim 22, wherein the active agent is administered in a copolymer or a gel coating over the surface.
 27. The method of claim 1, wherein said active compound has a structure of Formula A1:

wherein: each R is C1-12 alkyl; each A is, independently, hydrogen or an electron withdrawing group; M is metal selected from the group consisting of manganese, iron, copper, cobalt, nickel and zinc, and Z⁻ is a counterion.
 28. The method of claim 1, wherein said active compound has the formula:

wherein Z— is a counterion.
 29. The method of claim 1, wherein said active compound has a structure of Formula B1:

wherein: each R is C1-12 alkyl; each A is, independently, hydrogen or an electron withdrawing group; M is metal selected from the group consisting of manganese, iron, copper, cobalt, nickel and zinc, and Z— is a counterion.
 30. The method of claim 1, wherein said active compound has the structure:

wherein Z⁻ is a counterion.
 31. The method of claim 1, wherein said active compound has a structure of Formula C1:

wherein: each R is —(CH2)mCH2OX; m is 1 or 2; X is C1-12 alkyl; each A is, independently, hydrogen or an electron withdrawing group; M is metal selected from the group consisting of manganese, iron, copper, cobalt, nickel and zinc, and Z— is a counterion.
 32. The method of claim 1, wherein said active compound has the structure:

wherein Z⁻ is a counterion. 